Pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), have significant potential in tissue engineering and regenerative medicine due to the unlimited self-renewal ability and the capacity to differentiate into three-germ layers (1-3). In addition to PSCs' intrinsic properties, the extracellular PSC microenvironment comprising extracellular matrix (ECM) proteins as well as growth factors/cytokines also plays important roles in PSC development and function. In vivo, embryonic development is accompanied by the coordinated synthesis and secretion of ECM proteins, which are known to direct cell growth, survival, differentiation and morphogenesis (4,5). In vitro, cultured PSCs produce endogenous ECM proteins (i.e., fibronectin, basement membrane proteins) that regulate PSC fate through cell adhesion and/or binding with autocrine factors (i.e., leukemia inhibitory factor (LIF), Wnt, and Activin) (6-9). Thus, the derivation of ECMs from PSC cultures while preserving their distinct signaling capacities will greatly enhance their potential in cell delivery and tissue repair (10,11).
Stem cell-derived ECMs have been used to support in vitro cell expansion and differentiation as well as in vivo tissue regeneration (12-14). Tissue-specific ECMs derived from mesenchymal stem cells (MSCs) directed MSC lineage specification and augmented tissue regeneration by extending site-specific MSC retention (13-17). In addition, microcarriers made from MSC-derived ECMs promoted MSC adipogenesis and demonstrated in vivo compatibility, indicating the feasibility of their use in large scale cell production and cell/matrix delivery (14,15,18,19). The ECMs decellularized from ESC cultures provided a permissive microenvironment for tissue remodeling and fibroblast repopulation (11,12). The decellularized ESCs grown on Matrigel have also been shown to recapitulate the cellular and molecular milieu of the embryonic microenvironment and to sustain a balance between self-renewal and differentiation, preventing aberrant cell proliferation such as the aggressive cancer cells (20,21). Compared to the ECMs derived from adult stem cells or somatic tissues, the decellularized matrices from PSCs may have a broader spectrum of signaling capacity owing to their embryonic origin (6,10). The PSC-derived ECMs are free of embryonic DNA and thus have reduced risk of tumor formation, significantly improving their prospects in clinical applications.
In vitro, PSCs have been grown as undifferentiated monolayers or 3-D aggregates for expansion, or as embryoid bodies (EBs) differentiated into specific lineages (22-25). These organizations have been shown to affect the secretion of ECM proteins and autocrine factors, producing ECM microenvironment of distinct characteristics (20,26,27). For example, deposition of endogenous transforming growth factor-beta (TGF-β) inhibitor, Lefty, into ECMs has been reported for undifferentiated ESCs but not EBs, whereas the EBs displayed unique ECM-shell structure and different cytokine secretion profiles compared to undifferentiated ESCs (20,28-30). PSCs expanded as aggregates up-regulated E-cadherin expression and down-regulated Wnt signaling upon differentiation compared to PSCs cultured on 2-D substrate, suggesting the impact of intercellular interactions on cell signaling (26). Parallel to the organizational dependence, the characteristics of PSC-derived ECMs are also influenced by lineage specifications (6,23,29). For example, cerberus, a small antagonist of bone morphogenic protein (BMP), was detected in the secretome and ECMs of ESCs undergoing cardiac differentiation but not neural differentiation (6,28). Finally, the ECMs derived at the primitive or definitive stage of EBs exhibited different signaling capacities, suggesting the influence of developmental stage on ECM characteristics (10,11,28).